Drug addiction continues to remain a major public health concern in the United States. Addictive behavior results from changes in central nervous system signaling pathways that are modified after exposure to drugs of abuse. In particular, compounds such as cannabinoids and opiates that influence mood and pain perception are commonly associated with addictive behaviors. Many receptors regulating addiction are pharmacologically and biochemically well characterized, but some orphan receptors like GPR35 with homology to known receptors of abuse remain almost totally uncharacterized. The identification of small molecules capable of selectively inhibiting or activating orphans will provide enabling tools for elucidating novel molecular pathways underlying addictive behaviors. This proposal seeks to provide new compounds for characterizing the orphan G protein-coupled receptor GPR35. We have identified pamoic acid analogs as potent GPR35 agonists using in vitro assays and have found that pamoic acid induces antinociception. We propose a strategy to identify predominantly commercially available small molecules towards the objective of identifying a useful molecular probe for GPR35. Optimizing these novel compounds will allow the characterization of GPR35 biology in vitro and in animal models of pain. Our results suggest unexpected biological functions of pamoic acid and potential application for new drug development. PUBLIC HEALTH RELEVANCE: This proposal will provide tools for delineating the pharmacology of GPR35, potentially provide compounds for targeted therapeutics of pathways underlying pain, and clarify our understanding of GPR35 biology in vitro and in animal models of pain.